Serum Neutralizing Antibody Levels Research Articles

Recombinant Bovine Herpesvirus 4 (BoHV-4) Expressing Glycoprotein D of BoHV-1 Is Immunogenic and Elicits Serum-Neutralizing Antibodies against BoHV-1 in a Rabbit Model

Several biological characteristics of bovine herpesvirus 4 (BoHV-4) make it a good candidate as a gene delivery vector for vaccination purposes. These characteristics include little or no pathogenicity, unlikely oncogenicity, the capability to accommodate large amounts of foreign genetic material, the ability to infect several cell types coming from different animal species, and the ability to maintain transgene expression in both undifferentiated and differentiated cells. Starting from BoHV-4 cloned as a bacterial artificial chromosome (BAC), we used MuA transposase-mediated in vitro transposition to generate recombinant BoHV-4 expressing the immunodominant glycoprotein D (gD) of BoHV-1, one of the most important pathogens of cattle. Although a cis-acting element from woodchuck hepatitis virus (the woodchuck hepatitis virus posttranscriptional regulatory element [WPRE]) in the 3' end of the gD expression cassette was required for maximal gD expression from plasmids in transient transfection assays, this element was not necessary for efficient expression of gD from recombinant BoHV-4 genomes. BoHV-4 recombinants containing gD expression cassettes with or without the WPRE expressed gD at similarly high levels. Several cell lines originating from different animal species expressed gD when infected with BoHV-4 recombinants. When rabbits were immunized with one of the recombinants, high levels of serum neutralizing antibodies against BoHV-1 were generated. This work is one of the first demonstrations of the use BoHV-4 as a vector for vaccine purposes and may provide the basis for BoHV-1 vaccination of cattle with recombinant BoHV-4.

296. Transgene Expression and Neutralizing Antibody Production after Submandibular Gland Administration of AdhAQP1

Top of pageAbstract The treatment of patients with head and neck cancers typically includes ionizing radiation (IR). A significant consequence of this is the irreversible damage to salivary glands in the IR field. Without saliva, these patients suffer considerable morbidity, including oral infections, mucositis, dysphagia, as well as frank discomfort. Currently, there is no adequate treatment for this condition. Previously (Delporte et al, PNAS, 1997; Shan et al, Mol Ther, 2005), we showed that serotype 5 adenoviral (Ad5) -mediated transfer of the human aquaporin-1 (hAQP1) cDNA, via AdhAQP1, resulted in a restoration of near normal salivary flow to both rat submandibular and minipig parotid glands four months following IR. In preparation for clinical studies testing this strategy, we conducted a toxicology study with AdhAQP1 in Fischer 344 rats over a 13-week period. Herein, we describe the assessment of transgenic hAQP1 expression in targeted salivary glands, as well as the neutralizing antibody responses to vector delivery. A total of 100 animals per gender were studied. On day zero, animals received either vehicle (diluent buffer), or AdhAQP1 (2 |[times]| 10e8, 8 |[times]| 10e9 or 2 |[times]| 10e11 particle units; pu) via retroductal submandibular gland delivery. Five animals per gender and dosage group were sacrificed on days 3, 15, 29, 57 and 92. Protein from submandibular gland aqueous extracts (1000|[times]|g supernatants) was slot blotted onto nitrocellulose filters, and hAQP1 detected with a polyclonal antibody. Neutralizing antibodies were measured as the dilution of serum that resulted in 50% inhibition of the transduction of 293 cells with an Ad5 vector encoding luciferase, AdRSVLuc (MOI = 10). All vector-treated animals survived until scheduled sacrifice, and in general toxicological observations were comparable to findings previously reported with similar Ad5 vectors expressing different transgenes (O'Connell et al, J Oral Pathol Med, 2003; Zheng et al, Oral Dis, 2006). Transgene expression was readily seen in day 3 samples and was detectable in most high dose samples through the day 15-time point. Thereafter (days 29, 57, 92), slot blot results from vector-treated animals were not different from those of vehicle-treated animals. Additionally, on day 3, the expression of transgene was vector dose-dependent. None of the vehicle-treated rats showed the presence of neutralizing antibodies to Ad5 vectors in their sera. However, by day 29, four animals in the 2 |[times]| 10e8 pu vector dosage group exhibited low levels of serum neutralizing antibodies (2/gender; 1:4, 1:8, 1:8, 1:16), while all animals receiving 8|[times]|10e9 pu AdhAQP1 were positive for neutralizing antibodies (1:64 |[ndash]| 1:256; median 1:128). Similarly, all animals receiving 2|[times]|10e11 pu AdhAQP1 were positive for neutralizing antibodies on day 29, but with much higher titers (1:256 |[ndash]| 1:2048; median 1:1024). These results show that following AdhAQP1 administration to rat submandibular glands, hAQP1 protein is expressed, and anti-Ad5 neutralizing antibodies are produced, in a dose and time-dependent manner.

Generation of the transgenic potato expressing full-length spike protein of infectious bronchitis virus

To seek a new delivery system of vaccine for infectious bronchitis virus (IBV), transgenic potato expressing full-length spike (S) protein of IBV was produced and its immunogenicity in chickens was investigated. One to three copies of S gene of IBV were randomly and stably inserted into potato ( Solanum tuberrosum cv. Dongnong 303) genome by Agrobacterium tumefaciens-mediated transformation. Transcription and translation of S gene for IBV were confirmed by Northern blot and Western blot analyses in transgenic plantlets. Chickens immunized orally and intramuscularly with transgenic potato tubers expressing S protein generated the detectable levels of serum neutralizing antibodies and were protected against the challenge with the virulent IBV. In vitro secretion of interleukin 2 and T lymphocyte proliferation of spleen cells from the immunized chickens varied with the dose and the manner of vaccination with S protein derived from transgenic plants. The results indicated that S protein expressed in transgenic plants might be a new source for the production of Coronaviridae IBV vaccine.

Maternal antibody blocks humoral but not T cell responses to BVDV

Bovine viral diarrhoea virus (BVDV) contributes significantly to health-related economic losses in the beef and dairy industry. Antibodies of maternal origin can be protective against BVDV infection, however, calves with low titres of maternal antibody or that do not receive colostrum may be at risk for acute BVDV infection. Interference by high titres of maternal antibodies prevents the development of an antibody response following vaccination with either a killed or attenuated BVDV vaccine. However, the T cell mediated immune response to BVDV may be generated in the absence of a detectable serum neutralizing antibody response. Two trials were conducted to evaluate the potential to elicit T cell mediated immune responses to BVDV in calves with circulating maternal antibody to BVDV. In the first trial, calves with high levels of circulating maternal antibody to BVDV 1 and BVDV 2 were experimentally infected with BVDV 2 (strain 1373) at two to five weeks of age. The T-cell mediated immune responses of the experimentally infected calves and non-infected calves were monitored monthly until circulating maternal antibody was no longer detectable in either treatment group. Calves experimentally infected with BVDV developed BVDV specific CD4 +, CD8 +, and δ T cell responses while high levels of maternal antibody were circulating. A second challenge with BVDV 2 (strain 1373) was performed in the experimentally infected and control calves once maternal antibody could no longer be detected. Previous exposure to BVDV in the presence of maternal antibody protected calves from clinical signs of acute BVDV infection compared to the control calves. In the second trial, three groups of calves with circulating maternal antibody to BVDV were given either a modified live vaccine (MLV) containing BVDV 1 and BVDV 2, a killed vaccine containing BVDV 1 and BVDV 2, or no vaccine, at seven weeks of age. Serum neutralizing antibody levels and antigen specific T cell responses were monitored for 14 weeks following vaccination. Calves vaccinated with MLV BVDV developed BVDV 1 and BVDV 2 specific CD4 +T cell responses, and BVDV 2 specific γδ T cell responses, in the presence of maternal antibody. Vaccination with killed BVDV did not result in the generation of measurable antigen specific T cell immune responses. In this trial, a second vaccination was performed at 14 weeks to determine whether an anamnestic antibody response could be generated when calves were vaccinated in the presence of maternal antibody. Calves vaccinated with either a MLV or killed BVDV vaccine while they had maternal antibody developed an anamnestic antibody response to BVDV 2 upon subsequent vaccination. The results of these trials indicate that vaccinating young calves against BVD while maternal antibody is present may generate BVDV specific memory T and B cells. The data also demonstrated that seronegative calves with memory T and B cells specific for BVDV may be immune to challenge with virulent BVDV.

Correlates of immunity to respiratory syncytial virus (RSV) associated-hospitalization: establishment of minimum protective threshold levels of serum neutralizing antibodies

Objective: To determine if respiratory syncytial virus (RSV) specific, serum antibody titers correlate with protection against RSV associated-hospitalization at all ages. Design: Participants who were enrolled in a trial to determine the frequency of specific virus infections associated with hospitalization [J. Am. Med. Assoc. 283 (2000) 499] were included in our analysis if they were enrolled from July 1991 to June 1993, had a culture for virus isolation, and provided blood samples at hospitalization and 14–60 days later. RSV infection was defined by a positive culture and/or serology. Microneutralization, ELISA to the fusion (F) protein and Western blot were the serological assays that were used to determine correlates of immunity. Results: One hundred and seventy-five individuals, 1 month to 89 years old, out of 538 patients hospitalized with an acute respiratory infection met the criteria for analysis. RSV associated-hospitalization occurred in 11 (40.7%) of 27 infants (<1 year), 8 (38.1%) of 21 young children (1 to <5 years), and 15 (11.8%) of 127 children and adults (≥5 years). At the time of hospitalization, geometric mean neutralizing antibody titers (log 2) to RSV/A and RSV/B, and geometric mean binding antibody titer (log 2) to F protein were significantly higher in patients with non-RSV associated-hospitalization compared to those with RSV associated-hospitalization (RSV/A: 7.9 versus 6.1, P<0.001; RSV/B: 9.4 versus 7.3, P<0.001; ELISA-F, 13.9 versus 12.6, P=0.01). For every 1 log 2 increase in titer of neutralizing antibodies to RSV/A and RSV/B, and binding antibody to F protein there was a significant increase in the likelihood of not having an RSV associated-hospitalization by 22.3, 25, and 24.4% respectively. A minimal protective threshold titer of ≥6.0 (odds ratio 3.5; 95% CI 1.4–9.1) and ≥8.0 log 2 (odds ratio 2.9; 95% CI 1.1–7.7) against RSV associated-hospitalization was established for neutralizing antibodies to RSV/A and RSV/B; a threshold titer could not be established for binding antibody to F protein. Conclusion: Participants with naturally acquired serum neutralizing antibody levels at least equal to the minimal protective threshold titer were approximately three times more likely not to have an RSV associated-hospitalization. We speculate that achieving a minimal protective threshold antibody titer through active immunization will significantly reduce RSV associated-hospitalization among all ages.

Efficacy and safety studies of a recombinant chimeric respiratory syncytial virus FG glycoprotein vaccine in cotton rats.

Several formulations of a recombinant chimeric respiratory syncytial virus (RSV) vaccine consisting of the extramembrane domains of the F and G glycoproteins (FG) were tested in cotton rats to evaluate efficacy and safety. The FG vaccine was highly immunogenic, providing nearly complete resistance to pulmonary infection at doses as low as 25 ng in spite of inducing relatively low levels of serum neutralizing antibody at low vaccine doses. Upon RSV challenge animals primed with FG vaccine showed quite mild alveolitis and interstitial pneumonitis, which were eliminated by the addition of monophosphoryl lipid A to the formulation.

Limited breadth of the protective immunity elicited by simian immunodeficiency virus SIVmne gp160 vaccines in a combination immunization regimen.

We previously reported that immunization with recombinant simian immunodeficiency virus SIVmne envelope (gp160) vaccines protected macaques against an intravenous challenge by the cloned homologous virus, E11S. In this study, we confirmed this observation and found that the vaccines were effective not only against virus grown on human T-cell lines but also against virus grown on macaque peripheral blood mononuclear cells (PBMC). The breadth of protection, however, was limited. In three experiments, 3 of 10 animals challenged with the parental uncloned SIVmne were completely protected. Of the remaining animals, three were transiently virus positive and four were persistently positive after challenge, as were 10 nonimmunized control animals. Protection was not correlated with levels of serum-neutralizing antibodies against the homologous SIVmne or a related virus, SIVmac251. To gain further insight into the protective mechanism, we analyzed nucleotide sequences in the envelope region of the uncloned challenge virus and compared them with those present in the PBMC of infected animals. The majority (85%) of the uncloned challenge virus was homologous to the molecular clone from which the vaccines were made (E11S type). The remaining 15% contained conserved changes in the V1 region (variant types). Control animals infected with this uncloned virus had different proportions of the two genotypes, whereas three of four immunized but persistently infected animals had >99% of the variant types early after infection. These results indicate that the protective immunity elicited by recombinant gp160 vaccines is restricted primarily to the homologous virus and suggest the possibility that immune responses directed to the V1 region of the envelope protein play a role in protection.

Seroprevalence of Antibody Against Poliovirus in Inner-city Preschool Children

To assess susceptibility to poliomyelitis in selected inner-city preschool children in the United States and to estimate the contribution of secondary spread of live attenuated oral poliovirus vaccine virus to type-specific immunity. Cross-sectional seroprevalence study. Serum neutralizing antibody levels against poliovirus types 1, 2, and 3 were analyzed according to vaccination status, age, and other sociodemographic variables. Hospital and satellite clinics serving inner-city populations in Houston, Tex, and Detroit, Mich, 1990 to 1991. A total of 526 children aged 12 to 47 months seeking medical care were enrolled in the seroprevalence study; 144 children aged 12 to 35 months without a history of previous oral poliovirus vaccination were enrolled in the secondary spread study. Seropositive rates were similar in children in both cities, ranging from about 80% for types 1 and 3 in 12- to 23-month-old children to more than 90% in those aged 36 to 47 months. The most important predictor of seropositivity was the number of doses of oral poliovirus vaccine received (P < .01), with levels approximately 90% for all 3 serotypes among children who had received 3 or more doses. In children likely to have been unvaccinated, seropositive rates ranged from 9% to 18% for poliovirus types 1 and 3 and from 29% to 42% for type 2; secondary spread of vaccine virus appeared to have occurred among children who had previously received 1 dose or less but not those with 2 or more doses. Levels of immunity to poliovirus among inner-city preschoolers are high and may be predicted by the number of doses of oral poliovirus vaccine received. Secondary spread of the vaccine virus plays a modest role in increasing polio immunity in inner-city populations, especially against types 1 and 3. This role will decrease in importance if the recently attained high levels of immunization coverage in the United States are sustained and if the risk of importation of wild poliovirus continues to diminish.

Measles immunity and response to revaccination among secondary school children in Cumbria

The prevalence of antibody to measles virus in 759 children aged 11-18 years attending a secondary school in Cumbria was measured using a salivary IgG antibody capture assay. Serum IgG antibody levels were measured using a plaque reduction neutralization assay in subjects whose saliva was antibody negative. Vaccination histories were obtained from the child health computer and general practice record. A total of 662 pupils (87% of those tested) had detectable measles-specific IgG in saliva. Of the remaining 97, 82 provided blood samples and 29 had serum neutralizing antibody levels above 200 mIU/ml. Afer adjusting for non-participation rates, the proportion considered non-immune (no IgG in saliva and < or = 200 mIU/ml in serum) was 9% overall, ranging from 6% in vaccinated children to 20% in unvaccinated children. Measles-mumps-rubella vaccine was given to 50 children of whom 38 provided post-vaccination serum and 32 saliva samples. Thirty (79%) had a fourfold or greater rise in serum neutralizing antibody and 28 (88%) developed IgG antibody in saliva. Half of the children considered non-immune by antibody testing would have been overlooked in a selective vaccination programme targeted at those without a history of prior vaccination. A programme targeted at all school children should substantially reduce the proportion non-immune since a primary or booster response was achieved in three quarters of previously vaccinated children with low antibody levels and in all unvaccinated children. While it is feasible to screen a school-sized population for immunity to measles relatively quickly using a salivary IgG assay, a simple inexpensive field assay would need to be developed before salivary screening and selective vaccination could substitute for universal vaccination of populations at risk of measles outbreaks. The salivary IgG assay provided a sensitive measure of a booster response to vaccination.

A cotton rat model of effectors of immunity to respiratory syncytial virus other than serum antibody

A model for studying effectors of immunity to respiratory syncytial virus (RSV) was developed. Paris of inbred cotton rats (Sigmodon hispidus) were joined surgically using the technique of parabiosis. One week later, one animal of each pair was primed intranasally with a small volume of RSV suspension. Fourteen days after priming, both animals of each pair were bled for determination of serum neutralizing antibody titers, and challenged intranasally with a standard dose of RSV suspension. Single, unprimed cotton rats were challenged concomitantly and served as controls. Four days after challenge, all animals were sacrificed for virus titration of nasal tissues and lungs. Parabiosed cotton rats were surgically separated at varying intervals between priming and challenge (days 7, 9, 12, or 14 after priming) or were kept joined until sacrificed (day 18). Significant transfer of nasal and pulmonary immunity from primed to unprimed parabionts began 9 days after priming, gradually increasing through 18 days. Resistance to RSV challenge in spite of low levels of serum neutralizing antibody suggests that non-antibody immunologic mediators were responsible for the transferred immunity. Evidence is presented for three broad categories of RSV immunologic effectors: systemic, local with a transient systemic phase, and local without a systemic phase. These categories are now amenable to further study using the described model.

A subunit gIV vaccine, produced by transfected mammalian cells in culture, induces mucosal immunity against bovine herpesvirus-1 in cattle

A truncated version of bovine herpesvirus-1 (BHV-1) glycoprotein IV (tgIV) was produced in a novel, non-destructive expression system based upon regulation of gene expression by the bovine heat-shock protein 70A (hsp70) gene promoter in Madin Darby bovine kidney (MDBK) cells. In this system, up to 20 micrograms ml-1 of secreted tgIV, which is equivalent to the yield from 4 x 10(6) cells, was produced daily over a period of up to 18 days. Different doses of tgIV were injected intramuscularly into seronegative calves. Virus-neutralizing antibodies were induced by all doses of tgIV, both in the serum and in the nasal superficial mucosa. However, the low dose (2.3 micrograms) induced significantly (p < 0.05) lower antibody titres than the medium (7 micrograms) and high (21 micrograms) doses. The medium and high doses of tgIV conferred protection from BHV-1 infection, as demonstrated by a significant (p < 0.05) reduction in clinical signs of respiratory disease and virus shedding in the nasal secretions postchallenge. However, the 2.3 micrograms group, although partially protected, was not significantly (p > 0.05) different from the placebo group. This study demonstrated the potential of an intramuscularly administered tgIV subunit vaccine to induce mucosal immunity to BHV-1 using an economic protein production system and an acceptable vaccine formulation. In addition, a strong correlation was observed between neutralizing antibodies in the serum and nasal superficial mucosa, virus shedding and clinical disease. Thus, serum neutralizing antibody levels in tgIV-immunized animals may be a good prognosticator of protection from BHV-1 infection and disease.

Mucosal Immunity Induced by Enhanced-Potency Inactivated and Oral Polio Vaccines

Oral polio vaccine (OPV) is recommended for routine immunization in the United States in part because of its ability to induce intestinal and pharyngeal immunity to reinfection. Mucosal immunity produced by OPV and enhanced-potency inactivated polio vaccine (E-IPV) was compared by challenging vaccines with type 1 OPV. Fewer OPV (25%) than E-IPV (63%) vaccinees excreted OPV virus in stool after challenge. The mean stool virus titer was higher and the duration of shedding longer among E-IPV excreters. Only one E-IPV and three OPV vaccinees shed virus in the pharynx after challenge. Prechallenge serum neutralizing antibody levels were not statistically different among E-IPV vaccinees who did and did not shed virus; these levels were much higher than those of OPV vaccinees. Poliovirus-specific IgA levels in stool did not correlate with viral excretion. E-IPV was less effective than OPV in preventing and limiting intestinal infection, even though it induced higher postvaccination serum antibody levels.

Synthetic peptides of envelope proteins of two different strains of simian aids retrovirus (SRV-1 and SRV-2) represent unique antigenic determinants for serum neutralizing antibodies

There are at least three distinct serotypes of simian type D retrovirus (SRV) which exhibit extensive serological cross-reactivity, but no cross-reactivity exists at the level of serum neutralizing antibodies. Amino acid sequence analysis and hydrophobicity plots of SRV-1 and SRV-2 envelope proteins were compared in order to identify unique potential antigenic determinants to which respective neutralizing antibodies may be directed. Peptides representing residues 147–162 of SRV-1 and 96–102 of SRV-2 were synthesized and assessed for their immunoreactivity. Free peptide inhibition of strain-specific serum (rhesus) neutralizing antibodies to SRV-1 and SRV-2 was demonstrated using the SRV-1 147–162 peptide and the SRV-2 peptide, 96–102, respectively. Inhibition of serum neutralizing activity by these peptides was also strain-specific, showing no cross-inhibition. SRV-1 147–162 conjugated to a protein carrier and cross-linked to Sepharose beads specifically adsorbed neutralizing antibodies from SRV-1 immune rhesus sera. The antibodies eluted from the immunoadsorbent possessed SRV-1 neutralizing activity, but showed no effect on the infectivity of SRV-2. Peptide SRV-1 147–162 also exhibited the capacity to bind specifically with a mouse monoclonal antibody which neutralizes the infectivity of SRV-1. Mice immunized with a recombinant SRV-1 envelope protein or with whole, inactivated SRV-1 produced antibodies which bound the SRV-1 147–162 conjugate, but not the protein carrier itself. Mouse antibodies to the SRV-1 147–162 conjugate exhibited specific binding with both natlve SRV-1 and with recombinant SRV-1 envelope protein. These findings provide strong evidence that SRV-1 147–162 and SRV-2 96–102 constitute at least two unique antigenic determinants, or parts thereof, which participate in the strain-specific neutralizing antibody response. Moreover, the findings indicate that the SRV-1 neutralizing antibodies produced by monkeys and at least a certain population of neutralizing antibodies produced by mice recognize the same epitope of SRV-1.

Passive immunization protects the mouse eye from damage after herpes simplex virus infection by limiting spread of virus in the nervous system

Mice were treated with serum containing antibodies to herpes simplex virus type 1 (HSV-1) or normal serum, 1 day before inoculation on the cornea with HSV-1 strain McKrae. As expected, without passive immunization, mice developed high levels of serum neutralizing antibody. By contrast, in passively immunized animals, such antibody became undetectable by 29 days after inoculation of serum, in spite of the virus infection. There was no difference between passively immunized mice and those given normal serum in the duration of shedding of virus in tears and the duration and severity of corneal epithelial disease. However, non-immunized mice had a high incidence of mortality and developed disease of the iris, corneal stroma and lids, and their corneas became opaque and vascularized. In non-immunized animals, the timing of isolation of virus from nervous tissues and the sequence of appearance of virus antigens in ocular tissues indicate that the disease of deeper eye tissue was caused by virus spreading from the nervous system back to the eye. Restriction of such spread in passively immunized animals seems the likely explanation for their protection from death and severe ocular damage. Despite this restricted spread, passively immunized animals had a high incidence of latent infection in the ophthalmic part of the trigeminal ganglion. However, in comparison with mice given normal serum, there was a far lower incidence of such infection in the other two parts of this ganglion and in the superior cervical ganglion. Since passively immunized animals have a high incidence of latent infection in the ophthalmic part of the trigeminal ganglion and their eyes are normal, they will prove useful in studies involving induction of recurrent disease.

1074 SAFETY AND IMMUNOGENICITY OF A LIVE ATTENUATED RHESUS MONKEY ROTAVIRUS VACCINE

Rotavirus is a major worldwide cause of acute gastroenteritis in children. We recently evaluated a rhesus monkey rotavirus (RRV-1) which shares neutralization specificity with human sero-type 3, as a potential vaccine candidate. We administered RRV-1 (106 TCID50) or placebo orally to 40 adult volunteers who had low or undetectable levels of serum neutralizing antibody under double-blind conditions. Volunteers were evaluated daily for signs and symptoms of gastroenteritis, and stools were examined for RRV-1 by ELISA and tissue culture. Administration of RRV-1 did not induce illness. RRV-1 was shed in the stools of 11/21 (53%) vaccinees for 1 to 6 days without transmission of RRV-1 to other subjects, although one placebo recipient shed wild-type rotavirus from a community-acquired infection. Fourfold or greater serum neutralizing antibody rises to RRV-1 were noted in all vaccinees (21/21) and in 1/18 placebo recipients. Serum antibody rises were also demonstrated by immune adherence hemagglutination in 20/21 volunteers who received RRV-1 and in 1/18 who received placebo, and by complement fixation in 17/21 who received RRV-1 and 0/17 who received placebo. Transient asymptomatic serum ALT and AST rises were seen with similar frequencies in RRV-1 (3/21) and placebo (5/19) recipients. Thus, RRV-1 appears to be attenuated and highly immunogenic in young adults, and warrants additional study in children.

Mechanisms of immunity to respiratory syncytial virus in cotton rats.

Active immunity and maternally transmitted passive immunity to respiratory syncytial virus (RSV) were studied in cotton rats. Animals infected with respiratory syncytial virus developed complete resistance to pulmonary reinfection, which lasted at least 18 months. Nasal resistance was of shorter duration and began to diminish in 8 months. Pulmonary resistance was transferred by parabiosis, but nasal resistance was not. Adoptive transfer studies with fractionated convalescent blood showed that serum antibody, but not circulating lymphocytes, conferred pulmonary resistance. Immune females conferred antibody to their young prenatally and postnatally, with most of the antibody being transferred via colostrum and milk. Maternally transmitted immunity was more effective in the lungs than in the nose and was transient in both organs. Foster nursing experiments showed colostrum and milk to be the most important routes of immune transfer. Although resistance in infants generally correlated with serum neutralizing antibody levels, several exceptions to this correlation suggested that immune factors other than neutralizing antibody may also play an important role in maternal passive immunity.

Junin virus infection in genetically athymic mice.

The progression of Junin virus infection was studied in congenitally athymic mice. Immunocompetent littermates were used as infected controls. As expected, the latter developed lethal encephalitis, with viremia and considerable viral replication in the brain. The mortality rate was almost 100%; the few surviving controls exhibited high serum neutralizing antibody levels and a total absence of virus in blood and brain. In contrast, nude mice did not contract the disease; all survived with persistent viremia and virus in brain, but no serum neutralizing antibodies were detected. These results confirm previous research on thymectomized mice and those treated with anti-lymphocyte serum and tend to support the important role of cellular immunity in the pathogenesis of this viral disease.

A more sensitive technique for isolating infectious pancreatic necrosis virus from asymptomatic carrier rainbow trout, Salmo gairdneri Richardson

Abstract. An improved technique for isolating infectious pancreatic necrosis virus (IPNV) is described. It involves trypsinization of the suspect tissue, separation of the trypsinized cells and their co‐cultivation with indicator RTG‐2 cells. This co‐cultivation method is compared with homogenization, followed by membrane filtration, a procedure routinely employed by most disease diagnostic laboratories. From the two IPN‐infected farms studied, IPNV could be isolated by homogenization from only 50 and 66%, respectively, of the fish from which virus was detected by co‐cultivation. This difference in sensitivity can be explained in two ways, viz: (a) some of the virus is being retained by the membrane filter during filtration to remove contaminants following homogenization; (b) the presence of neutralizing factors in the tissue homogenate are neutralizing the virus and thus no cytopathic effect ensues. No correlation between serum neutralizing antibody levels and the differences in sensitivity of the two methods was found. The co‐cultivation method is as practical as, and certainly no more time‐consuming than, the homogenization technique.

The Importance of an Intact Complement Pathway in Recovery from a Primary Viral Infection: Influenza in Decomplemented and in C5-Deficient Mice

Abstract To investigate the role of complement (C) in the recovery from a primary viral infection in vivo, the course of influenza infection in decomplemented BALB/c mice and in C5-deficient B10.D2/OSN mice was examined. BALB/c male mice decomplemented with cobra venom factor demonstrated a prolonged viral infection, an increased degree of pulmonary consolidation, and a higher rate of morbidity and mortality than did nondecomplemented mice for each dose of virus administered. This deleterious effect occurred in mice decomplemented before infection and immunologic induction, as well as in mice decomplemented after infection and immunologic induction, provided the C3 levels remained depressed at the time of viral clearance (days 7 to 9). The time of appearance and the levels of serum neutralizing antibodies and viral-specific cytotoxic T cells were equivalent in decomplemented and nondecomplemented BALB/c mice infected with influenza. Similarly, C5-deficient B10.D2/OSN male mice, infected with influenza, demonstrated a prolonged viral infection, an increased degree of pulmonary consolidation and a higher rate of mortality than did C5-sufficient B10.D2/NSN agematched mice for each dose of virus administered. These results suggest than an intact complement pathway plays an adjunctive role in host recovery from primary influenza infection.

Serum Neutralizing Antibody Development and Host Resistance in Chickens Exposed to Marek’s Disease Infection

The activity of serum neutralizing (SN) antibodies in both resistant and susceptible line chickens was quantitated by SN tests in vitro cell cultures against JM virus. The levels of SN antibodies were higher in genetically resistant chickens than in susceptible ones after they were inoculated with JM strain of Marek’s disease virus at day-old. The relation between the production of SN antibodies and the resistance to Marek’s disease and oncogenicity was positively correlated.